Delafield's
Alum Hematoxylin

Ingredient	Amount	Function
Hematoxylin	6 g	Dye
Ammonium alum	90 g	Mordant
Distilled water	600 mL	Solvent
95% ethanol	200 mL	Solvent
Glycerol	150 mL	Stabiliser

 

Compounding procedure
Dissolve the hematoxylin in 50 mL ethanol.
Dissolve the alum in the water.
Add the two solutions.
Leave, loosely stoppered, in a warm, sunlit place for one week.
Filter, then add the glycerol and the rest of the ethanol.
Leave, loosely stoppered, in a warm, sunlit place for 3 months.
Filter and store in a tightly stoppered container in the dark.
This solution is stable for years.

Method

1. Bring sections to water with xylene and ethanol.
2. Place into the staining solution for 20 minutes.
3. Rinse well with water.
4. Differentiate with acid ethanol if necessary.
5. Rinse with water and blue.
6. Rinse well with water.
7. Counterstain if desired.
8. Dehydrate with ethanol, clear with xylene and mount with a resinous medium.

Expected results

• Nuclei  –  blue
• Background  –  as counterstain or unstained

Notes

1. The solution may be chemically ripened by adding 0.5 g sodium iodate, but such solutions are considered to be inferior in longevity.
2. Like many strong regressive formulations, cartilage, cement lines and mucins may be blue.
3. Acid ethanol is 0.5% - 1% hydrochloric acid in 70% ethanol.
4. Blueing is done with alkaline solutions such as hard tap water, Scott’s tap water substitute, 0.1% ammonia water, 1% aqueous sodium acetate, 0.5% aqueous lithium carbonate etc.

 

Reference
Culling, C.F.A., Allison, R.T. and Barr, W.T.
Cellular Pathology Technique, Ed.4.
Butterworth, London, UK.

Drury, R.A.B. and Wallington, E.A., (1980)
Carleton's histological technique Ed. 5
Oxford University Press, Oxford, UK.

Bancroft, J.D. and Stevens A. (1982)
Theory and practice of histological techniques Ed. 2
Churchill Livingstone, Edinburgh & London, UK.